[[4-[[2-(4-cyclohexylphenoxy)ethyl]ethylamino]-2-methylphenyl]methylene]malononitrile

Long-term toxicity to aquatic invertebrates

Administrative data

Endpoint:

long-term toxicity to aquatic invertebrates

Type of information:

experimental study

Adequacy of study:

key study

Reliability:

1 (reliable without restriction)

Rationale for reliability incl. deficiencies:

guideline study

Data source

Reference

Reference Type:

study report

Title:

Unnamed

Year:

2008

Report date:

2008

Materials and methods

Test guideline

Test guideline 1

Qualifier:

equivalent or similar to guideline

Guideline:

OECD Guideline 211 (Daphnia magna Reproduction Test)

Test guideline 2

Qualifier:

according to guideline

Guideline:

EU Method C.20 (Daphnia magna Reproduction Test)

GLP compliance:

yes (incl. QA statement)

Test material

Test material information

Constituent 1

Reference substance name:

[[4-[[2-(4-cyclohexylphenoxy)ethyl]ethylamino]-2-methylphenyl]methylene]malononitrile

EC Number:

258-964-5

EC Name:

[[4-[[2-(4-cyclohexylphenoxy)ethyl]ethylamino]-2-methylphenyl]methylene]malononitrile

Cas Number:

54079-53-7

Molecular formula:

C27H31N3O

IUPAC Name:

2-[(4-{[2-(4-cyclohexylphenoxy)ethyl](ethyl)amino}-2-methylphenyl)methylidene]propanedinitrile

Sampling and analysis

Analytical monitoring:

yes

Details on sampling:

Sampling schedule:
- Control: on day 0, 7 and 14 in the freshly prepared media; on day 2, 9 and 16 in the old media
- Test concentrations: on day 0, 7 and 14 in the freshly prepared media; on day 2, 9 and 16 in the old media

Test solutions

Vehicle:

no

Details on test solutions:

To accelerate the solution procedure, a higher amount of the water solubility of Solvent Yelow 179 (< 0.0007 mg/L) was taken (2 mg) and added directly to 2 litres of dilution water, treated for 1 h in an ultrasonic bath and then stirred for 24 h on a magnetic stirrer. Finally undissolved particles of the test item were removed by filtration using a folded filter of pore size 7-12 μm.

Test organisms

Test organisms (species):

Daphnia magna

Details on test organisms:

- Source: Strain of Bundesgesundheitsamt Berlin
- Maintenance and Acclimatisation: A population of parthenogenetic females of synchronized age structure has been maintained for more than 15 years in the test facility under constant temperature conditions (20 +/- 1 °C) at a 16:8 hour light-dark photoperiod (illumination: < 1000 lux). The culture water (so-called 'M4 medium') was partly renewed once a week. The Daphnia were exclusively fed with unicellular green algae (Desmodesmus subspicatus) 'ad libitum'. Mortalities of parent Daphnia during the culture period were recorded daily in a semi-quantitative way. The neonates were separated from their parent Daphnia by filtration prior to the acute test.
- Culture and dilution water: Reconstituted water (so-called 'M4 medium')

Study design

Test type:

semi-static

Water media type:

freshwater

Limit test:

yes

Total exposure duration:

21 d

Test conditions

Hardness:

16.1 - 16.8 °dH (= 287 - 300 mg/L CaCO3)

Test temperature:

20 +-1 °C

pH:

7.8 - 8.0

Dissolved oxygen:

8.5 - 9.2 mg/L

Salinity:

n.a.

Conductivity:

no data

Nominal and measured concentrations:

Nominal concentration: 1 mg/L; After 16 days the values were below or at the limit of quantification (0.002 mg/L); Recovery rates ranged from 0.6–3.1 % of the nominal values in the freshly prepared media and from 0.6–3.2 % in the media after 48 hours, 72 hours of exposure, respectively.

Details on test conditions:

EXPOSURE CONDITIONS:
- Test vessels: 150 mL glass beakers holding 1 neonate in 100 mL of test medium
- Experimental design: 1 test concentration plus 1 control; 1 neonate per vessel, 10 replicates per concentration/control semi-static system (renewal of the test media on Monday, Wednesday and Friday) respectively.
- Method of initiation: neonates were placed in prepared media
- Photoperiod: 16 h light: 8 h dark
- Light intensity: not exceeding 800 lux
- Aeration: none
- Food source: Desmodesmus subspicatus
- Feeding rate: day 0-7: 0.1 mg C / Daphnia / day; day 8-21: 0.2 mg C / Daphnia / day
- Criteria of effects: The criterion of adverse effects used in this study was the item-induced inhibitory effect on the parthenogenetic reproduction rate and the mobility behaviour/mortality rate of parent Daphnia, all effects being assessed at least three times a week.

Reference substance (positive control):

no

Results and discussion

Effect concentrations

Effect concentrations 1

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Effect concentrations 2

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

> 1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

reproduction

Effect concentrations 3

Duration:

21 d

Dose descriptor:

NOEC

Effect conc.:

>= 1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Effect concentrations 4

Duration:

21 d

Dose descriptor:

LOEC

Effect conc.:

> 1 mg/L

Nominal / measured:

nominal

Conc. based on:

test mat.

Basis for effect:

mortality

Details on results:

The results are expressed in terms of nominal concentrations. Recovery rates ranged from 0.6 – 3.1 % of the nominal values in the freshly prepared media and from 0.6 – 3.2 % in the media after 48 hours.

Applicant's summary and conclusion

Validity criteria fulfilled:

yes

Remarks:

The mortality rate in the controls did not exceed 20% by the end of the test; Living offspring produced per parent Daphnia surviving at the end of the test was ≥ 60 in the controls; DOC > 3 mg/L; pH: 6-9; water hardness > 140 mg/L (as CaCO3).

Conclusions:

No adverse effects were observed at the limit of water solubility (21d-NOEC(reproduction): ≥ 1mg/L, 21d-LOEC(reproduction): > 1 mg/L, 21d-NOEC(mortality): ≥ 1 mg/L, 21d-LOEC(mortality): > 1 mg/L). The results are expressed in terms of nominal concentrations. Recovery rates ranged from 0.6–3.1 % of the nominal values in the freshly prepared media and from 0.6–3.2 % in the media after 48 hours, 72 hours of exposure, respectively.

Executive summary:

The study was conducted in accordance with EEC Methods for Determination of Ecotoxicity Annex to Directive 97/548/EEC Part C, Method 20 'Daphnia magna Reproduction Test'' (adopted: 2001), which equals to the OECD Test guideline 211 (Daphnia magna Reproduction Test; adopted: Sept. 21, 1998). Young female Daphnia, aged less than 24 hours at the start of the test, were exposed to a limit test concentration of nominally 1 mg/L of Solvent Yellow 179 dissolved in water. Auxiliaries used to prepare the test media were an ultrasonic bath, a magnetic stirrer and folded filters. After a 21d-exposure period, the total number of living offspring produced per parent animal alive at the end of the test was assessed. The reproductive output of the Daphnia exposed to the test item was compared to that of the control in order to determine the Lowest Observed Effect Concentration (LOEC) and the No Observed Effect Concentration (NOEC) according to DUNNETT (1955, 1964). Additionally, the survival of the parent daphnia and the time to production of first brood was determined. The following values were determined (mg/L): 21d-NOEC(reproduction): ≥ 1, 21d-LOEC(reproduction): > 1, 21d-NOEC(mortality): ≥ 1, 21d-LOEC(mortality): > 1. The 1st brood was observed on day 8. Thus, no adverse effects were observed at the limit of water solubility. This toxicity study is classified as acceptable and satisfies the guideline requirements for the chronic daphnia study.